MIT Department of Biology
7.02 Experimental Biology & Communication, Spring 2005
RDM Day 3 Interpretation Questions
1. On Day 4, you see the following number of colonies on each of your transformation
plates. (These plates are labeled as the transformations are labeled on page 18 of your
RDM manual.)
Plate #
1
2
3
4
5
6
# of colonies
77
22
8
~700
212
7
Interpret these results by including the answers to the questions: Do you think that you
were successful in the goal of this experiment? Did you have any contamination? Did
your CIP treatment from Day 2 work completely? Explain your answers.
2. You have a mutant strain of cells that contains the lacZ gene in the 7.02 mini-Tn10
inserted into one of three genes: mrk, nat, or irm. You also have wild-type cells of the
same strain with no lacZ insertion. Your UROP Professor asks you to design an
experiment to identify which gene received the insertion. You design very similar sets of
primers to those you used in 7.02 on RDM Day 3. You run the following PCR reactions
and load a gel with the following lanes:
Lane
1
2
3
4
5
6
7
8
9
10
11
12
13
What’s loaded in that lane
1 kb ladder
PCR rxn on control DNA with mrk For and Rev primers
PCR rxn on control DNA with nat For and Rev primers
PCR rxn on control DNA with irm For and Rev primers
PCR rxn on control DNA with mrk For and lacZ Rev primers
PCR rxn on control DNA with nat For and lacZ Rev primers
PCR rxn on control DNA with irm For and lacZ Rev primers
PCR rxn on mutant DNA with mrk For and Rev primers
PCR rxn on mutant DNA with nat For and Rev primers
PCR rxn on mutant DNA with irm For and Rev primers
PCR rxn on mutant DNA with mrk For and lacZ Rev primers
PCR rxn on mutant DNA with nat For and lacZ Rev primers
PCR rxn on mutant DNA with irm For and lacZ Rev primers
You correctly determine that lacZ inserted into nat. Draw what your gel looked like,
showing any bands that you saw in each lane. (You saw a band of 1.2kb in lane 2, a band
of 1.8 kb in lane 3, and a band of 2.7 kb in lane 4.) Please label each band with its correct
approximate size.